Nanopore adaptive sampling for mitogenome sequencing and bloodmeal identification in hematophagous insects

Abstract

Blood-feeding insects are important vectors for an array of zoonotic pathogens. Despite significant research focused on well-documented insect vectors of One Health importance, resources for molecular species identification of a large number of hematophagous arthropods are limited. Advancements in next-generation sequencing technologies provide opportunities for targeting mitochondrial genomes of blood-feeding insects, as well as their bloodmeal hosts. This dual approach holds great promise for elucidating complex disease transmission pathways and enhancing the molecular resources for the identification of cryptic insect species. To this end, we leveraged the newly developed Oxford Nanopore Adaptive Sampling (NAS) pipeline to dually sequence the mitogenomes of hematophagous insects and their bloodmeals. Using NAS, we sequenced the entire mitogenomes of Aedes vexans, Culex restuans, Culex territans, and Chrysops niger and successfully identified bloodmeal hosts of Chrysops niger, Culex restuans, and Aedes trivittatus. We show that NAS has the utility to simultaneously molecularly identify blood-feeding insects and characterize disease transmission pathways through bloodmeal host identification. Moreover, our data indicate NAS can facilitate a wide array of molecular systematic studies through novel ‘phylogenetic capture’ methods. We conclude the NAS approach has great potential for informing global One Health initiatives centered on the mitigation of vector-borne disease through dual vector and bloodmeal identification.

Utilisation of Oxford Nanopore sequencing to generate six complete gastropod mitochondrial genomes as part of a biodiversity curriculum

“…Given sufficient sequencing coverage, we found basecalling with high accuracy (hac) mode was able to produce mitogenomes with complete genes without premature stop codons. In addition to enriching for mitochondrial DNA experimentally 15 , 54 , the use of adaptive sampling recently developed in ONT allows preselected sequences to be enriched during the sequencing process 55 – 57 . This approach has proven successful in obtaining full mitogenomes of endangered animals by enriching host genomic DNA from fecal samples 56 .…”

Genome Enrichment of Rare, Unknown Species from Complicated Microbiome by Nanopore Selective Sequencing

“…The newly designed Readfish toolkit eliminates the need for complex signal mapping algorithms, and exploits existing ONT tools to provide a robust toolkit for designing and controlling selective sequencing experiments 10 . Until now, the application of RU is principally limited to the elimination of known host species 9, 10, 11 or the enrichment of known targets such as mitogenomes of blood-feeding insects 12, 13 .…”